Protein kinases are enzymes dedicated to transfer phosphate groups from ATP to a substrate protein. Phosphorylation of the targeted proteins results in a functional change of their activity and can also modify the association with other proteins, trafficking and subcellular localization. It is estimated that up to 30% of all proteins can be modified by kinases. For this reason, kinases are key regulators of the majority of cellular pathways, especially those involved in signal transduction. Phosphorylation is a mode of transmission of information on biomolecular level.
There are receptor protein kinases, which are located in cellular membranes, and non-receptor protein kinases, which are located in the cytoplasm.
Cyclin dependent protein kinases (CDKs) are non-receptor kinases that require cyclin for their activity. CDKs comprise a family of Ser/Thr kinases divided into two groups, including the cell cycle CDKs, which orchestrate cell cycle progression, and the transcriptional CDKs, which contribute to transcriptional regulation [Malumbres et al., Nat Rev Cancer 9: 153-166, (2009); Sausville, Trends Mol Med 8: S32-S37, (2002)]. The first group encompasses core components of the cell cycle machinery, including cyclin D-dependent kinases 4 and 6, as well as cyclin E-CDK2 complexes, which sequentially phosphorylate the retinoblastoma protein, Rb, to facilitate the G1/S transition. Cyclin A-dependent kinases 2 and 1 are required for orderly S phase progression, whereas cyclin BCDK1 complexes control the G2/M transition and participate in mitotic progression [Pines, Semin Cancer Biol 5: 305-313, (1994)]. The functional activation of cell cycle CDKs depends in part on the formation of heterodimeric cyclin-CDK complexes, which may be modulated by association with endogenous Cip/Kip or INK4 inhibitors [Sherr et al., Genes Dev 13: 1501-1512, (1999)]. CDKs are also regulated by phosphorylation, including positive events directed by CDK-activating kinase (CAK, cyclin H/CDK7/MAT1) and negative phosphorylation events [Morgan, Nature 374: 131-134, (1995)].
The transcriptional CDKs, including cyclin H-CDK7, cyclin C-CDK8 and cyclin T-CDK9 (P-TEFb), promote initiation and elongation of nascent RNA transcripts by phosphorylating the carboxy-terminal domain (CTD) of RNA polymerase II. CDK8 is a part of the Mediator complex that functions as a transcriptional coactivator in all eukaryotes. In addition, two other kinase components of this complex CDK11 and CDK19 (which is structurally similar to CDK8) were described [Drogat et al. Cell Reports 2: 1-9, (2012)]. CDK8 functions as an oncoprotein, especially in colorectal cancers where it regulates activity of β-catenin, and there is considerable interest in developing drugs specifically targeting the CDK8 kinase activity [Firestein et al., Nature 7212: 547-551, (2008)].
CDK8 resides on a region of Chr. 13 that is known to undergo chromosomal gain in 40-60% of colorectal cancers. Moreover, high CDK8 expression was detected in 70% tumors by immunohistochemistry [Adler et al., Cancer Res. 72: 2129-2139, (2008)]. Colorectal cancer cells that express elevated CDK8 levels are highly dependent on its expression for proliferation [Firestein et al., Nature 7212: 547-551, (2008)]. CDK8 was required to promote rapid tumor growth as well as maintain the CRC tumors in an undifferentiated state. CDK8 expression induced focus formation, anchorage-independent colony growth and tumor formation in immunodeficient animals [Adler et al., Cancer Res. 72: 2129-2139, (2008)].
CDK8 levels are also elevated in response to loss of the histone variant macroH2A (mH2A) [Kapoor et al. Nature 468(7327): 1105-1109, (2011)]. Loss of histone isoform mH2A promotes malignant phenotype of melanoma cells. Tumor promoting functions of mH2A are at least partially mediated by up-regulation of CDK8. Knockdown of CDK8 was able to suppress the enhanced proliferation of melanoma cells induced by mH2A loss in vitro and in vivo.
CDK8 is also involved in secretory activity of senescent cells in response to chemotherapy. Selective inhibition of CDK8 and CDK19 repressed expression of certain cytokines and growth factors which are released in response to chemotherapy treatment and stimulate tumor growth [Porter et al., PNAS (34) 109: 13799-13804, (2012)]. The role of CDK8 in expression of proinflammatory cytokines such as TNFα and IL6, upon stimulation with exogenous and endogenous factors, such as LPS and other TLR agonists, was so far not reported in the literature to the best of our knowledge. Therefore CDK8 can be considered as a novel, emerging target in the treatment of autoimmune and inflammatory disorders.
Suppression of CDK8 kinase is also an attractive strategy for targeting colorectal cancers, including these resistant to anti-EGFR therapies due to activating mutations in KRAS and BRAF downstream in the pathway [Donner et al., Nat Struct Mol Biol. 17: 194-201, (2010)].
In contrast CDK8 deficiency in cultured “normal” metazoan cells did not affect cell viability [Westerling et al., Nature. 382:638-42 (1996)]. Hence, inhibitors of CDK8 are considered as promising agents for cancer.
High expression of CDK8 significantly increased colon cancer-specific mortality [Firestein et al., Int J Cancer 126(12): 2863-2873, (2010)] and decreased duration of relapse-free survival in patients with breast and ovarian cancer [Porter et al., PNAS (34) 109: 13799-13804, (2012)].
The majority of small molecule inhibitors block kinases by binding to the ATP binding site, which is highly conserved, especially in the family of CDKs. Most of the known CDK inhibitors are, however, rather unselective and display undesired side effects. Small molecules which selectively target the transcriptional kinase CDK8 are thus desirable when treating e.g. cancer, autoimmune and inflammatory diseases.